The present invention relates to a tryptase inhibitor comprising, as an active ingredient, one or more kinds of substances selected from guanidinoaliphatic acid derivatives, guanidinobenzoic acid derivatives, guanidinophenol derivatives, amidinophenol derivatives, and nontoxic salts, acid addition salts or hydrates thereof, and to novel guanidinophenol derivatives and nontoxic salt, acid addition salt or hydrate thereof. More specifically, the present invention relates to a tryptase inhibitor comprising, as an active ingredient, one or more kinds of substances selected from guanidinoaliphatic acid derivatives, guanidinobenzoic acid derivatives, guanidinophenol derivatives and amidinophenol derivatives represented by the formulae (I) to (IV) 
(in the formulae, all the symbols represent the same meanings as hereafter defined), or nontoxic salts, acid addition salts or hydrates thereof, and to pharmaceuticals comprising, as an active ingredient, novel guanidinophenol derivatives represented by the formula (V) 
(in the formulae, all the symbols represent the same meanings as hereafter defined), or nontoxic salts, acid addition salts or hydrates thereof.
Tryptase is an intracellular trypsin-type protease that was isolated and purified from the human lung by Smith, et al. in 1984 (see T. J. Smith, M. W. Houglandi D. A. Johnson, J. Biol. Chem., 25, 11046 (1984)).
Tryptase is thought to belong to the trypsin-type protease since (1) tryptase cleaves the C-terminal of basic amino acids (especially arginine) as well as trypsin, (2) it decomposes common trypsin artificial substrates, and (3) its protease activity is inhibited by the trypsin inhibitors.
Tryptase is, however, categorized into a family having different properties from trypsin on the basis that (1) tryptase cannot decompose Arg-MCA (arginine-methylcoumarinamide) and Arg-bNA (arginine-xcex2-naphthylamide) which are artificial substrates highly decomposed by trypsin, (2) it highly decomposes Boc-Ile-Gly-Arg-MCA(t-butoxycarbonyl-isoleucine-glycine-arginine-methylcoumarinamide) which is an artificial substrate of blood coagulation factor Xa that is hard to be decomposed by trypsin, and (3) it limitedly decomposes prothrombin to produce thrombin while trypsin nonlimitedly decomposes most prothrombin. Additionally, tryptase is completely different from the other protease (e.g., plasmin, thrombin, kallikrein, elastase. etc.) in the structure, localizing cells, physiological functions and so on.
Tryptase is known as it exists in the following cells, and however, the structure thereof varies depending upon the localizing cell.
(1) Tryptase in Mast Cell (Tryptase M)
Tryptase M localizes in the soluble state in the typical (connective tissue) mast cell, atypical (membrane) mast cell and the histamine granules of basocytes. Human tryptase M was isolated and purified by Schwartz, et al. (see L. B. Schwartz, R. A. Lewis, K. F. Ansten, J. Biol. Chem., 25, 11939 (1981)). This enzyme is a heterotetramer having a molecular weight of 144,000 composed of two molecules of subunit having a molecular weight of 37,000 and two having that of 35,000.
(2) Tryptase in T4 Lymphocyte (Tryptases TL1, TL2 and TL3)
Tryptases TL1, TL2 and TL3 were isolated and purified from the bulk-cultured cells of the Molt 4 clone 8 strain, which is one of the T4 lymphocytes, by Kido, Katunuma, et al. (see H. Kido, A. Fukutomi, N. Katunuma, J. Biol. Chem., 2, 21979 (1991)). Tryptases TL1 and TL2 were purified from the cell membrane, and Tryptase TL3 was purified from the Golgi body.
(3) Tryptase in Clara Cells (Tryptase Clara)
Tryptase Clara was isolated and purified from the bronchial membrane secretory cell (Clara cell) by Kido, Katsunuma, et al. (see H. Kido, Y. Yokogoshi, K. Sakai, M. Tashiro, Y. Kishino, A. Fukutomi, N. Katunuma, J. Biol. Chem., 267, 13573 (1992)). The molecular weight of this enzyme is 30,000 by means of SDS-PAGE (SDS-polyacrylamide gel electrophoresis) in the presence of a reducing agent, or 180,000 by gel filtration.
The physiological functions of the above-mentioned tryptase have been reported as follows:
(1) decomposition of fibrinogen (see J. Immunol., 135, 2762 (1985)),
(2) decomposition of polymer kininogen and fibrin (see J. Immunol., 135, 2762 (1985)),
(3) activation of single-stranded urokinase type plasminogen activator (see J. Biol. Chem., 269, 9416 (1994)),
(4) formation of complement C3a from C3 (see J. Immunol., 130, 1891 (1983)),
(5) decomposition of VIP (vaso active intestinal peptide) and CGRP (calcitonin gene-related peptide) (see Allergy, 27, 90 (1990), J. Pharmacol. Exp. Ther., 244, 133 (1988), and J. Pharmac. and Exp. Ther., 248, 947 (1989)),
(6) release of IL-8 from the bronchial epithelial cell and stimulation to express ICAM-1 (see J. Immunol., 15, 275 (1996)),
(7) decomposition of fibronectin (see J. Cell. Biochem., 50, 337 (1992)).
(8) stimulation to proliferate fibroblast (see J. Crin. Invest., 88, 493 (1991)),
(9) decomposition of collagen IV,
(10) activation of promatrix metalloprotease 3 (proMMP-3) (J. Crin. Invest., 84, 1657 (1989)),
(11) activation of stromelysin, and
(12) decomposition of 72 kD gelatomase.
Studies on the relation of tryptase to condition of disorder have been reported as follows.
(1) Tryptase at high level exists in the washing water of the bronchus of asthma patients.
(2) The tryptase released from the mast cell in the site of pulmonary fibrosis induces inflammation reaction mediated by various mediators.
(3) The tryptase released from the mast cell in the site of chronic rheumatism enhances collagenase activity.
(4) The tryptase in the process of infection of Sendai virus elevates the ability of membrane fusion and infection of the virus.
Tryptase inhibitors are thought to be useful for prevention and/or treatment of various diseases caused by tryptase. In consideration of the relation between the physiological function of tryptase and condition of disorder, the diseases are exemplified by asthma, pulmonary fibrosis, interstitial pneumonitis, nephritis, hepatic fibrosis, hepatitis, cirrhosis, scleroderma, psoriasis, atopic dermatitis, chronic rheumatism, influenza, Crohn""s disease, inflammatory intestinal diseases, ulcerative colitis, nasal allergy, atherosclerosis, etc.
Recently, some studies on the tryptase inhibitors of nonpeptide compounds having an amidino or guanidino group have been reported (see J. Stuerzebecher, et. al., Biol. Chem. Hoppe-Seyler, 37, 1025 (1992), G. H. Caughey, et. al., J. Pharmacol. Exp. Ther. 2, 676 (1993), C. H. Kam, et al., Arch. Biochem. Biophys., 316, 808 (1995), U.S. Pat. No. 4,845,242, U.S. Pat. No. 5,089,634, U.S. Pat. No. 5,324,648, and PCT Publication No. 9,427,958).
In order to find tryptase inhibitors from the points of view above, the present inventors intensively studied to find that guanidinoaliphatic acid derivatives represented by the formula (I), guanidinobenzoic acid derivatives represented by the formula (II), guanidinophenol derivatives represented by the formula (III) and amidinophenol derivatives represented by the formula (IV) have potent tryptase inhibitory activity. Thus, the present invention is completed.
Further, it was found that novel guanidinophenol derivatives represented by the formula (V) have potent tryptase inhibitory activity.
Moreover, it was found that the novel guanidinophenol derivatives represented by the formula (V) have potent antagonistic inhibitory activity on a variety of proteinases (e.g., trypsin, plasmin, thrombin and kallikrein), phospholipase A2 (PLA2) and/or leukotriene B4 (LTB4).
That is, the present invention relates to
(1) a tryptase inhibitor comprising, as an active ingredient, guanidinoaliphatic acid derivatives represented by the formula (I) 
xe2x80x83(in the formula, R1 represents a hydrogen atom, a halogen atom, a nitro group, an alkyl group, an alkoxyl group, a carboxyl group or an alkoxycarbonyl group, and n represents an integer of 3 to 6), or nontoxic salts, acid addition salts or hydrates thereof;
(2) a tryptase inhibitor comprising, as an active ingredient, guanidinobenzoic acid derivatives represented by the formula (II) 
xe2x80x83(in the formula, R2 represents a phenyl group, a naphthyl group, a substituted phenyl group or a substituted naphthyl group, and
R3 represents one of various substituting groups), or nontoxic salts, acid addition salts or hydrates thereof;
(3) a tryptase inhibitor comprising, as an active ingredient, guanidinophenol derivatives represented by the formula (III) 
xe2x80x83(in the formula, R4 and R5 represent various substituting groups), or nontoxic salts, acid addition salts or hydrates thereof;
(4) a tryptase inhibitor comprising, as an active ingredient, amidinophenol derivatives represented by the formula (IV) 
xe2x80x83(in the formula, R6 and R7 represent various substituting groups), or nontoxic salts, acid addition salts or hydrates thereof;
(5) guanidinophenol derivatives represented by the formula (V) 
xe2x80x83[in the formula, A represents a group of the following formula: 
(in the group, R9 and R10 each represents independently a hydrogen atom or a C1-4 alkyl group),
R8 represents a group selected from the following formulae: 
(in the groups, R11, R12 and R13 each represents independently
(1) a hydrogen atom,
(2) a phenyl group,
(3) a C1-4 alkyl group substituted by a phenyl group,
(4) a C1-10 alkyl group,
(5) a C1-10 alkoxyl group,
(6) a C2-10 alkenyl group having 1 to 3 double bonds,
(7) a C2-10 alkynyl group having 1 to 2 triple bonds,
(8) a group of formula: R15xe2x80x94COXR16 
(in the group, R15 represents a single bond or a C1-8 alkylene group, X represents an oxygen atom or an NH-group, and
R16 represents a hydrogen atom, a C1-4 alkyl group, a phenyl group or a C1-4 alkyl group substituted by a pheny group), or
(9) a C3-7 cycloalkyl group; 
xe2x80x83represents a 4-7 membered monocyclic heteroring containing 1 to 2 nitrogen atoms,
R14 represents a hydrogen atom, a C1-4 alkyl group substituted by a phenyl group or a group of formula: COOR17 (in the group,
R17 represents a hydrogen atom, a C1-4 alkyl group or a C1-4 alkyl group substituted by a phenyl group);
provided that R11, R12 and R13 do not represent simultaneously hydrogen atoms, and when at least one group among R11, R12 and
R13 represents a group having a t-butoxycarbonyl group, the other groups do not represent groups having a carboxyl group.] or nontoxic salts, acid addition salts or hydrates thereof;
(6) methods of preparing the guanidinophenol derivatives represented by the formula (V), and nontoxic salts, acid addition salts or hydrates thereof;
(7) pharmaceuticals comprising, as an active ingredient, the guanidinophenol derivatives represented by the formula (V), or nontoxic salts, acid addition salts or hydrates thereof;
(8) agents for prevention and/or treatment of asthma, pulmonary fibrosis, hepatitis, interstitial pneumonitis, nephritis, hepatic fibrosis, hepatitis, cirrhosis, scleroderma, psoriasis, atopicdermatitis, chronicrheumatism, influenza, Crohn""s disease, inflammatory intestinal diseases, ulcerative colitis, nasal allergy, atherosclerosis which comprise, as an active ingredient, the guanidinoaliphatic acid derivatives represented by the formula (I), the guanidinobenzoic acid derivatives represented by the formula (II), the guanidinophenol derivatives represented by the formula (III), the amidinophenol derivatives represented by the formula (IV), or nontoxic salts, acid addition salts or hydrates thereof; and
(9) agents for prevention and/or treatment of chronic rheumatism, inflammatory intestinal diseases, psoriasis, stomach diseases induced by non-steroidal anti-inflammatory drugs, adult respiratory distress syndrome, myocardial infarction, allergic rhinitis, hemodialysis-inducing neutropenia, late asthma, various inflammatory diseases, allergic diseases, disseminated intravascular coagulation, pancreatitis, severe stages thereof and multiple organ failure which comprise, as an active ingredient, the guanidinophenol derivatives represented by the formula (V), or nontoxic salts, acid addition salts or hydrates thereof.
In the present invention, all isomers are included unless otherwise noted. For instance, alkyl group, alkoxyl group, alkylene group, alkenylne group and alkynylene group include straight and branched ones, and the alkenylene group includes E- and Z-body and EZ-mixture based on the double bond. The isomers generated from the asymmetric carbon atoms are also included when there is branched alkyl group, alkoxyl group, alkylene group, alkenylene group or alkynylene group.
In the formula (V), the C1-4 alkyl group represented by R9, R10, R16 and R17 means a methyl, ethyl, propyl, and butyl group, and these isomers.
In the formula (V), the C1-10 alkyl group represented by R11, R12 and R13 means a methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, and decyl group, and these isomers.
In the formula (V), the C1-10 alkoxyl group represented by R11, R12 and R13 means a methoxy, ethoxy, propyloxy, butyloxy, pentyloxy, hexyloxy, heptyloxy, octyloxy, nonyloxy, and decyloxy group, and these isomers.
In the formula (V), the C1-4 alkyl group substituted by a phenyl group represented by R10, R12, R13, R14, R16 and R17 means a methyl, ethyl, propyl, and butyl group substituted by one phenyl group and these isomers.
In the formula (V), the C2-10 alkenyl group having 1 to 3 double bonds represented by R11, R12 and R13 means an ethenyl, propenyl, butenyl, pentenyl, hexenyl, heptenyl, octenyl, nonenyl, decenyl, butadienyl, pentadienyl, hexadienyl, heptadienyl, octadienyl, nonadienyl, decadienyl, hexatrienyl, heptatrienyl, octatrienyl, nonatrienyl, and decatrienyl group, and these isomers.
In the formula (V), the C2-10 alkynyl group having 1 to 2 triple bonds represented by R11, R12 and R13 means an ethynyl, propynyl, butynyl, pentynyl, hexynyl, heptynyl, octynyl, nonynyl, decynyl, butadiynyl, pentadiynyl, hexadiynyl, heptadiynyl, octadiynyl, nonadiynyl, and decadiynyl group, and these isomers.
In the formula (V), the C3-7 cycloalkyl group represented by R11, R12 and R13 means a cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and cycloheptyl group.
In the formula (V), the C1-8 alkylene group represented by R15 means a methylene, ethylene, trimethylene, tetra-methylene, pentamethylene, hexamethylene, heptamethylene, and octamethylene group, and these isomers.
In the formula (V), the 4-7 membered monocyclic heteroring containing 1 to 2 nitrogen atoms represented by 
means a pyrrole, pyrrolidine, imidazole, imidazolidine, pyridine, piperidine, pyrazine, piperazine, and pyrimidine ring, etc.
Although each substance used as the active ingredient in the present invention is well-known, detailed description is given below.
The preferred compounds include guanidinoalphatic acid derivatives represented by the formula (I) 
(in the formula, R1 represents a hydrogen atom, a halogen, a nitro, alkyl, alkoxyl, carboxyl or alkoxycarbonyl group, and n represents an integer of 3 to 6), and nontoxic salts, acid addition salts or hydrates thereof.
The examples of the guanidinoaliphatic acid derivatives represented by the formula (I) which are included in the present invention are the compounds described in JP-B-47-21977, JP-B-50-2494, JP-B-45-33175, JP-B-49-2107, JP-A-48-29732, JP-A-49-24917, JP-A-51-75042 and JP-A-54-76532.
The more preferred compound includes compound 1: 
p-ethoxycarbonylphenyl 6-guanidinohexanoate, and nontoxic salts, acid addition salts and hydrates thereof.
The preferred compounds include guanidinobenzoic acid derivatives represented by the formula (II) 
(in the formula, R2 represents a phenyl group, a naphthyl group, a substituted phenyl group or a substituted naphthyl group, and
R3 represents one of various substituting groups), and nontoxic salts, acid addition salts or hydrates thereof.
The examples of the guanidinobenzoic acid derivatives represented by the formula (II) which are encompassed in the present invention are the compounds described in JP-A-48-29732 and 49-24917 (U.S. Pat. No. 3,824,267), JP-A-49-11842, JP-A-50-4038, JP-A-50-69035, JP-A-51-16631, JP-A-52-89640 (U.S. Pat. No. 4,021,472), JP-A-53-15412, JP-A-53-147044, JP-A-54-70241 and 55-55154 (U.S. Pat. No. 4,224,342), JP-A-55-115865 and 55-115863 (U.S. Pat. No. 4,283,418), JP-A-56-34662 (U.S. Pat. No. 4,310,533), JP-A-62-111963 and 63-165357 (EP-A-222608). JP-A-55-100356, JP-A-56-110664, JP-A-57-53454, JP-A-57-142956, JP-A-57-142957, JP-A-57-179146, JP-A-58-41855, JP-A-58-49358, JP-A-61-286361, JP-A-61-286362, JP-A-62-103058, JP-A-62-155253, and GB-A-2083818 and 2095239.
The more preferred compound includes compound 2: 
N,N-ddimethylcarbamoylmethyl p-(p-guanidinobenzoyloxy)phenylacetate, compound 3: 
p-(p-guanidinobenzoyloxy)phenylacetic acid, compound 4: 
6-amidinonaphtho-2-yl p-guanidinobenzoate, and nontoxic salts, acid addition salts and hydrates thereof.
The preferred compounds include guanidinophenol derivatives represented by the formula (III) 
(in the formula, R4 and R5 represent various substituting groups), and nontoxic salts, acid addition salts or hydrates thereof.
The examples of the guanidinophenol derivatives represented by the formula (III) which are encompassed in the present invention are the compounds described in JP-A-5-286922 and Chem. Pharm. Bull., 32, 4466-4477 (1984), and the compounds represented by the above-mentioned the formula (V).
The more preferred compound includes compound 5: 
p-(p-guanidinophenoxycarbonyl)cinnamic acid N-phenyl-N-ethoxycarbonylmethylamide, compound 6: 
p-(p-guanidinophenoxycarbonyl)-xcex1-methylcinnamic acid N-phenyl-N-ethoxycarbonylmethylamlde, compound 7: 
p-(p-guanidinophenoxycarbonyl)cinnamic acid N-(2-ethoxycarbonyl)piperldinylamide, compound 8: 
p-(p-guanidinophenoxycarbonyl)cinnamic acid N-(3-ethoxycarbonyl)piperidinylamide, compound 9: 
p-(p-guanidinophenoxycarbonyl)cinnamic acid N-(4-ethoxycarbonyl)piperidinylamide, compound 10: 
p-(p-guanidinophenoxycarbonyl)cinnamic acid N-(2-(S)-ethoxycarbonyl)pyrrolidinylamide, compound 11: 
p-(p-guanidinophenoxycarbonyl)cinnamic acid N-(2-(R)-ethoxycarbonyl)pyrrolidinylamide, compound 12: 
p-(p-guanidinophenoxycarbonyl)cinnamic acid N-(1-(S)-ethoxycarbonyl-3-ethoxycarbonyl)propylamide, compound 13: 
p-(p-guanidinophenoxycarbonyl)-xcex1-methylcinnamic acid N-allyl-N-ethoxycarbonylmethylamide, compound 14: 
p-(p-guanidinophenoxycarbonyl)benzoic acid N-phenyl-N-ethoxycarbonylmethylamide, compound 15: 
m-(p-guanidinophenoxycarbonyl)benzenesulfonamide, and nontoxic salts, acid addition salts and hydrates thereof.
The preferred compounds include amidinophenol derivatives represented by the formula (IV): 
(in the formula, R6 and R7 represent various substituting groups), and nontoxic salt, acid addition salt or hydrate thereof.
The examples of the amidinophenol derivatives represented by the formula (IV) which are included in the present invention are the compounds described in JP-A-58-41855, Japanese Patent Application No. Hei 5-252178 (EP-A-588655), JP-A-7-206801 (EP-A-656349) and Japanese Patent Application No. Hei 7-263599 (EP-A-703216).
The more preferred compound includes compound 16: 
p-(p-amidinophenoxycarbonyl)-xcex1-methylcinnamic acid N-allyl-N-ethoxycarbonylmethylamide, compound 17: 
p-(p-amidinophenoxycarbonyl)-xcex1-methylcinnamic acid N-allyl-N-carboxylmethylamide, compound 18: 
p-(p-amidinophenoxycarbonyl)-xcex1-methylcinnamic acid N-propargyl-N-ethoxycarbonylmethylamide, compound 19: 
p-(p-amidinophenoxycarbonyl)cinnamic acid N-(4-phenyl-methyl)piperadinylamide, compound 20: 
p-(p-amidinophenoxycarbonyl)phenylpropionic acid N-allyl-N-ethoxycarbonylmethylamide, compound 21: 
p-(p-amidinophenoxycarbonyl)-xcex1-methylcinnamic acid N-(1-(S)-ethoxycarbonyl-2-ethoxycarbonylethylthio)ethylamide, compound 22: 
p-(p-amidlnophenoxycarbonyl)-xcex1-methylcinnamic acid N-ethoxycarbonylmethyl-N-hexylamide, compound 23: 
p-(4-amidino-2-methoxycarbonylphenoxycarbonyl)-xcex1-methylcinnamic acid N-allyl-N-ethoxycarbonylmethylamide, compound 24: 
p-(p-amidinophenoxycarbonyl) -xcex1-methylcinnamic acid N-(1,1-bis(ethoxycarbonylmethyl)methyl-N-cyclohexylamide, compound 25: 
p-(p-amidinophenoxycarbonyl)-xcex1-methylcinnamic acid N-2-ethoxycarbonylethyl-N-3-methoxypropylamide, and nontoxic salts. acid addition salts and hydrates thereof.
In the present invention, each of the active ingredients can be used alone, or two or more kinds of the active ingredients can be incorporated into a preparation.
[Salts]
The compounds of the present invention described in the present specification may be converted into nontoxic salts or acid addition salts by means of the known methods.
The preferable salts are nontoxic and water-soluble ones. Suitable salts include salts of alkali metal (such as potassium and sodium), salts of alkaline-earth metal (such as calcium and magnesium), ammonium salts, and salts of pharmaceutically acceptable organic amine (such as tetramethylammonium, triethylamine, methylamine, dimethylamine, cyclopentylamine, benzylamine, phenethylamine, piperidine, monoethanolamine, diethanolamine, tris(hydroxymethyl)aminomethane, lysine, arginine, and N-methyl-D-glucamine).
The suitable acid addition salts are nontoxic and water-soluble ones. Suitable acid addition salts include, for instance, inorganic salts such as hydrochloride, hydrobromide, hydrolodide, sulfate, phosphate and nitrate, or organic salts such as acetate, lactate, tartarate, benzoate, citrate, methanesulfonate, ethanesulfonate, benzenesulfonate, toluenesulfonate, isethionate, glucuronate and gluconate, and the preferable ones are methanesulfonate salts.
The compounds of the present invention described in the present specification, or nontoxic salts or acid addition salts thereof may be converted into hydrate by means of the known methods.
The novel guanidinophenol derivatives encompassed in the present invention are the guanidinophenol derivatives represented by the formula (V): 
(in the formula, all the symbols represent the same meanings as the above). or nontoxic salts, acid addition salts or hydrates thereof.
The preferred compounds include the above-mentioned compounds 5 to 13, and nontoxic salts, acid addition salts or hydrates thereof.
[Preparation of the Compounds of the Present Invention]
Among the compounds of the present invention, the guanidinoaliphatic acid derivatives represented by the formula (I), the guanidinobenzoic acid derivatives represented by the formula (II), the guanidinophenol derivatives represented by the formula (III) and the amidinophenol derivatives represented by the formula (IV) each can be prepared by the methods described in the above-mentioned specifications.
Among the compounds of the present invention, the guanidinophenol derivatives represented by the formula (V) can be prepared by the following method.
Among the compounds of the present invention represented by the formula (V), the groups represented by R11, R12, R13 and R14 in the group R8 of which each represents the group not containing a carboxyl (hereinafter referred to as COOH) group and a t-butoxycarbonyl (hereinafter referred to as COO-t-Bu) group, that is, those represented by the formula (Va): 
(in the formula, A represents the same meaning as the above, and R8a represents the same meaning as R8 above; provided that R11, R12, R13 and R14 in R18 each represents the group not containing a COOH group and a COO-t-Bu group) are prepared by allowing a compound represented by the formula (VIa): 
(in the formula, A and R8a represent the same meanings as the above) and p-amidinophenol to esterification reaction.
The esterification reaction is known and exemplified by
(1) a method using acid halide,
(2) a method using mixed acid anhydride,
(3) a method using a condensing agent, etc.
The specific descriptions of these methods are as follows.
(1) The method using acid halide is carried out, for instance, by allowing carboxylic acid to react with acid halide (such as oxalyl chloride, thionyl chloride, etc.) in an inert organic solvent (such as chloroform, methylene chloride, diethyl ether, and tetrahydrofuran) or without solvent at a temperature between xe2x88x9220xc2x0 C. and the reflux temperature, and allowing the resulting acid halide to react with alcohol in the presence of tertiary amine (such as pyridine, triethylamine, dimethylaniline, and dimethylaminopyridine) in an inert organic solvent (such as chloroform, methylene chloride, diethyl ether, and tetrahydrofuran) at a temperature between 0 and 40xc2x0 C.
(2) The method using mixed acid anhydride is carried out, for instance, by allowing carboxylic acid to react with acid halide (such as pivaloyl chloride, tosyl chloride, and mesyl chloride) or an acid derivative (such as ethyl chloroformate, isobutyl chloroformate, etc.) in the presence of tertiary amine (such as pyridine, tiethylamine, dimethylaniline, and dimethylaminopyridine) in an inert organic solvent (such as chloroform, methylene chloride, diethyl ether, and tetrahydrofuran) or without solvent at a temperature between 0 and 40xc2x0 C., and allowing the resulting mixed acid anhydride to react with the corresponding alcohol in an inert organic solvent (such as chloroform, methylene chloride, diethyl ether, and tetrahydrofuran) at a temperature between 0 and 40xc2x0 C.
(3) The method using a condensing agent (such as 1,3-dicyclohexylcarbodiimide(DCC), 1-ethyl-3-[3-(dimethylamino) propyl]carbodiimide(EDC), 2-chloro-1-methylpyridinium iodide, etc.) is carried out, for instance, by allowing carboxylic acid to react with alcohol in an inert organic solvent (such as chloroform, methylene chloride, dimethylformamide, and diethyl ether) or without solvent, using or without using tertiary amine (such as pyridine, triethylamine, dimethylaniline, and dimethylaminopyridine), and using a condensing agent at a temperature between 0 and 40xc2x0 C.
These reactions of (1), (2) and (3) are desirably carried out under an inert gas (such as argon and nitrogen) atmosphere under an anhydrous condition.
Among the compounds represented by the formula (V), those of which at least one of the groups represented by R11, R12 and R13 in the group R8 represents a group having a COO-t-Bu group and the other groups each represents a group not having a COOH group, or R14 represents a COO-t-Bu group, that is, those represented by the formula (Vb): 
(in the formula, A represents the same meaning as the above, and R8b represents the same meaning as R8 above; provided that at least one of R11, R12 and R13 in R8 represents a group having a COO-t-Bu group and the others each represents a group not having a COOH group or R14 represents a COO-t-Bu group) are prepared by allowing a compound represented by the formula (VIb): 
(in the formula, all the symbols represent the same meanings as the above) and a compound represented by the formula (VIb): 
(in the formulae, R11b, R12b, R13b and R14b each represents the same meaning as R11, R12, R13 and R14, respectively; provided that at least one of R11b, R12b and R13b represents a group having a COO-t-Bu group, and the other groups each represents a group not having a COOH group, or R14b represents a COO-t-Bu group) to amidation reaction. This amidation reaction is performed in the same way as the above-mentioned methods for esterification (provided that the amine of formula (VIIb) is used instead of the amidinophenol).
Among the compounds represented by the formula (V), those of which at least one of the groups represented by R11, R12 and R13 in the group R8 represents a group having a COOH group and the other groups each represents a group not having a COO-t-Bu group, or R14 represents a COOH group, that is, those represented by the formula (Vc): 
(in the formula, A represents the same meaning as the above, and R8c represents the same meaning as R8 above; provided that at least one of R11, R12 and R13 in R8 represents a group containing a COOH group and the others each represents a group not containing a COO-t-Bu group, or R14 represents a COOH group) are prepared by hydrolyzing the COO-t-Bu group of the compounds represented by the formula (Vb) which is prepared by the above-mentioned method.
The hydrolysis of the t-butyl ester group is performed, for instance, in organic acid (such as trifluoroacetic acid) or inorganic acid (such as hydrochloric acid) or a mixture thereof in an inert organic solvent (such as methylene chloride, chloroform, methanol, dioxane, ethyl acetate, and anisole) at a temperature between 0 and 90xc2x0 C.
The compounds represented by the formulae (VIa) and (VIb) can be prepared by the known methods, the methods described in Japanese Patent Application No. Hei 5-252178 (EP-A-588655), and the similar methods as those described in Japanese Patent Application No. Hei 5-252178 (EP-A-588655).
In each reaction in the present specification, the reaction product can be purified by common purification means, for instance, distillation under normal or reduced pressure, high performance liquid chromatography using silica gel or magnesium silicate, thin layer chromatography, or column chromatography or washing, recrystallization, etc. Purification may be performed after each reaction or after some reactions.
The other starting materials and each reagent are known of themselves or can be prepared by the known methods.
[Effect of the Invention]
As described above, the tryptase inhibitors are thought to be useful for prevention and/or treatment of asthma, pulmonary fibrosis, interstitial pneumonitis, nephritis, hepatic fibrosis, hepatitis, cirrhosis, scleroderma, psoriasis, atopic dermatitis, chronic rheumatism, influenza, Crohn""s disease, inflammatory intestinal diseases, ulcerative colitis, nasal allergy, atherosclerosis, etc. Accordingly, the compounds of the present invention represented by the formulae (I), (II), (III) and (IV) which have tryptase inhibitory activity can be used in animals including human, especially in human, as agents for prevention and/or treatment of asthma, pulmonary fibrosis, interstitial pneumonitis, nephritis, hepatic fibrosis, hepatitis, cirrhosis, scleroderma, psoriasis, atopic dermatitis, chronic rheumatism, influenza, Crohn""s disease, inflammatory intestinal diseases, ulcerative colitis, nasal allergy, atherosclerosis, etc.
Moreover, the compounds of the present invention represented by the formula (V) are useful for prevention and/or treatment of diseases inhibiting the antagonism of a variety of proteinases (e.g., trypsin, plasmin, thrombin and kallikrein), phospholipase A2 (PLA2) and/or leukotriene B4 (LTB4) other than inhibiting tryptase (e.g., chronic rheumatism, inflammatory intestinal diseases, psoriasis, stomach diseases induced by non-steroidal anti-inflammatory drugs, adult respiratory distress syndrome, myocardial infarction, allergic rhinitis, hemodialysis-inducing neutropenia, late asthma, various inflammatory diseases, allergic diseases, disseminated intravascular coagulation, pancreatitis, severe stages thereof and multiple organ failure).
[Toxicity]
Extremely low toxicity has been confirmed in each of the active ingredients and nontoxic salts and nontoxic acid addition salts thereof encompassed in the present invention. For instance, the LD50 of methanesulfonate of compound 16 was 117 mg/kg when intravenously administered to male mice. Accordingly, any of the active substances encompassed in the present invention can be considered to be adequately safe and suitable for use as pharmaceuticals.
[Application to Pharmaceuticals]
The substances described in the present invention are usually administered systemically or locally, orally or parenterally when used for the above-mentioned purpose.
The dosage, however, depends upon the age, body weight, symptom, therapeutic effect, administering way, treatment time, etc., normally, oral administration with the dosage ranging between 1 and 1000 mg an adult is made once to a few times a day, or parenteral administration (preferably intravenous administration) with the dosage ranging between 1 and 100 mg an adult is made once to a few times a day, or continuous administration is intravenously made for 1 to 24 hours a day.
As described above, the dosage alters depending upon the various conditions, and therefore, less dosage than the above may be adequate or more may be necessary.
The compounds of the present Invention are administered as solid compositions, liquid compositions and the other compositions for oral administration, and as injections, external preparations, suppositories, etc. for parenteral administration.
The solid compositions for oral administration include tablets, pills, capsules, powders, granules, etc.
The capsules include hard capsules and soft ones.
In such solid compositions, one or more active substances are mixed with at least one inert diluent, e.g., lactose, mannitol, mannite, glucose, hydroxypropylcellulose, microcrystalline cellulose, starch, polyvinyl pyrrolidone and magnesium metasilicate aluminate. The compositions may contain additives except the inert diluent, e.g., tablet lubricants such as magnesium stearate, disintegrants such as fibrous calcium glycolate, and solution adjuvant such as glutamic acid and aspartic acid, in accordance with the conventional way. The tablets and pills may be coated, if necessary, with film of gastrically soluble or enteric substances such as white sugar, gelatin, hydroxypropylcellulose and hydroxypropylmethyl-cellulose phthalate, or with two or more layers. Additionally, capsules of absorbable substances such as gelatin are also included.
The liquid compositions for oral administration include pharmaceutically acceptable emulsions, solutions, syrups, elixirs, etc. In such liquid compositions, one or more active substances are contained ininert diluents generally used (e.g. purified water and ethanol). These compositions may contain adjuvants such as penetrants and suspending agents, sweetening agents, flavoring agents and antiseptics in addition to the inert diluents.
The other compositions for oral administration include sprays that contain one or more active substances and are formulated by the known methods in themselves. These compositions may contain stabilizers to add stability and isotonicity such as sodium hydrogensulfite, and tonicity agents such as sodium chloride, sodium citrate or citric acid. Methods of preparing sprays are described in detail in, e.g., U.S. Pat. Nos. 2,868,691 and 3,095,355.
The injections for parenteral administration according to the present invention include sterile aqueous or nonaqueous solutions, suspensions and emulsions. The aqueous solution and suspensions include, e.g., distilled water for injection and physiological saline. The nonaqueous solutions and suspensions are exemplified by propylene glycol, polyethylene glycol, vegetable oil such as olive oil, alcohol such as ethanol, POLYSORBATE 80 (registered trademark). These compositions may further contain additives such as antiseptics, penetrants, emulsifiers, dispersers, stabilizers, solution adjuvants (e.g., glutamic acid and aspartic acid). These are sterilized by filtration through bacterial filter, addition of bactericide or irradiation. These may be prepared as sterile solid compositions and used with dissolving in sterilized or sterile distilled water for injection or other solvents at use.
The other compositions for parenteral administration include external solutions, ointments, endermic liniments, suppositories for rectal administration and pessaries for vaginal administration which contain one or more active substances and are formulated by the conventional methods.